Project Summary Developmental exposure to inorganic arsenic has been associated with several adverse health effects, including reduction in cognitive performance and increased rates of psychiatric disorders. Altered programming of the glucocorticoid receptor (GR) could account for many of the long-lasting consequences of arsenic. While the impact of arsenic exposure on the epigenome has been studied in the context of cancer research, the influence of this toxicant in the developing brain, particularly as it relates to epigenetics, is not well understood. In our previous work, we found that male offspring exposed to 50 ppb arsenic prenatally (PAE) had decreased expression of both GR protein and mRNA in brain. Female PAE mice were resistant to arsenic induced changes in GR protein despite elevations in GR (Nr3c1) mRNA. In assessing epigenetic mechanisms, we found PAE affected levels of histone3 lysine4 trimethylation (H3K4me3) as well as several noncoding RNAs (miRs and lncRNA) in a sex-dependent manner. The goal of this competing renewal is to demonstrate that changes in transcriptional, posttranscriptional and posttranslational regulation result in PAE sex-dependent effects at a molecular, physiological and behavioral level. Specific Aim 1 will test the hypothesis that PAE alters histone modifications that control the transcription of stress-related genes during fetal development in a sex-specific manner. We will assess histone posttranslational modifications (HPTMs), H3K4 me3/ H3K27 me3, histone writers and erasers, and associated genes by western and sequential ChIP-qPCR techniques. We will confirm a functional link by pharmacologically altering specific histone marks at embryonic day 12.5 (E12.5). Specific Aim 2 will test the hypothesis that PAE alters the posttranscriptional regulation of stress-related mRNA levels through micro-RNAs (miRs) during fetal development in a sex- specific manner. These experiments will assess miR expression using qPCR and miR-RIP to target the GR signaling system. We will confirm a functional link using LNA microRNA mimics and LNA-antimiR at E12.5. Specific Aim 3 will test the hypothesis that PAE alters the posttranslational regulation of stress- related proteins through long noncoding RNA (lncRNA) during fetal development in a sex- specific manner. These experiments will assess growth arrest-specific 5 (Gas-5) expression at E12-18. We will confirm a functional link to the GR system using LNA-GAPmer and LNA?antimir approaches. Specific Aim 4 will test the mechanistic link between PAE-induced transcriptional, posttranscriptional and/or posttranslational changes and the functional outcomes in the adult animal. These experiments will assess the impact of interventions identified in Aims 1-3 on arsenic-induced deficits: corticosterone response, learning and depression.